Proj 2: Design and screening for immunogens to efficiently elicit anti-HIV-1 bNAbs PI: Pamela J. Bjorkman An effective HIV vaccine likely requires induction of Abs against HIV-1 Env that neutralize across the majority of circulating viral strains (broadly neutralizing antibodies; bNAbs). Because administered bNAbs are protective against HIV-1 infection in animal models, it is widely believed that a vaccine that elicits bNAbs would prevent infection. The fact that vaccine efforts have not yet succeeded preventing HIV-1 infection and/or eliciting bNAbs may relate to the observation that inferred germline (iGL) forms of bNAbs only rarely bind to Env proteins that are the targets of bNAbs. The germline targeting approach to vaccine design involves engineering an iGL-binding Env protein that preferentially activates bNAb precursors and selects productive somatic hypermutations (SHMs) during affinity maturation. Resulting memory B cells are then boosted by sequential immunogens to shepherd bNAb production by selecting additional productive SHMs. Drs. Bjorkman and Nussenzweig propose to apply this approach to target two classes of HIV-1 bNAb: a class related to PGT121 that binds to the base of the V3 variable loop and interacts with the N332gp120 glycan (V3/N332 bNAbs), and IOMA-like bNAbs, a new class of CD4-mimetic CD4 binding site (CD4bs) bNAbs derived from the VH1-2 germline gene segment. The V3/N332 and IOMA classes were chosen because (i) V3/N332 Abs are among the most potent of bNAbs, are commonly found in HIV-infected individuals who develop bNAbs, and immunogen studies will be complementary to human clinical trials evaluating passive delivery of the V3/N332 bNAb 10-1074 to HIV-1?infected patients, (ii) IOMA's relatively low number of SHMs and its normal-length CDRL3 suggest it may be more easily elicited than VRC01-class VH1-2?derived CD4bs bNAbs that are heavily somatically mutated and contain rare 5-residue CDRL3s, and (iii) immunogen design will be facilitated by the recent structure of a natively-glycosylated Env trimer bound to the V3/N332 bNAb 10-1074 and to IOMA. With the goal of creating SOSIP-based Env trimers to target iGLs and shepherd maturation of V3/N332 and IOMA-like bNAbs, Dr. Bjorkman proposes a highly collaborative project with Dr. Nussenzweig to (1) Solve structures of iGL?immunogen complexes to aid in structure-based immunogen design and library screens to select immunogens, (2) Use structure-based design combined with library screening to identify SOSIP trimers that bind V3/N332 iGLs with high affinity, (3) Use structural information to guide construction of a yeast display library to find rare variants that bind IOMA iGL with high affinity, (4) Combine results from Aims 2 and 3 to create SOSIP-based immunogens that bind iGLs of both bNAbs and evaluate them in wildtype mice and in mice produced by Dr. Nussenzweig to carry germline-reverted versions of IOMA and 10-1074. These aims will be repeated using information from new structures, library screens, and screens for Abs isolated by Dr. Nussenzweig from HIV-nave individuals and from sequentially-immunized animals.